Brigitte Schoener

Intracellular Oxidation-Reduction States in Vivo The microfluorometry of pyridine nucleotide gives a continuous measurement of the oxidation state

It now appears to be possible to continuously record changes in intracellular oxidation-reduction levels in terms of the fluorescence of reduced pyridine nucleotide in mitochondria of various tissues and organs in situ. Studies of kidney and brain cortex in the rat show that changes in fluorescence are not measurably affected by the presence of oxyhemoglobin. Nitrogen, sulfide, cyanide, and carbon monoxide cause increases in fluorescence to very nearly the same levels, and the increases are attributed to larger reduction of mitochondrial diphosphopyridine nucleotide. Amytal at a low blood concentration causes increased reduction in the kidney cortex, and at a high blood concentration, in the brain cortex. The qualitative response of the pyridine nucleotide to low oxygen concentrations shows the brain to be more sensitive than the kidney. The first measurable increase in pyridine nucleotide reduction observed on the brain occurs at a concentration of inspired oxygen of 8 percent. Breathing stops when the percentage increase of pyridine nucleotide reduction on the brain reaches about 90; at this point the percentage increase for the kidney is only about 30. This difference corresponds roughly to a tenfold difference in oxygen tension. Half-maximal increase in pyridine nucleotide reduction on the brain occurs at a concentration of inspired oxygen of about 4 percent and corresponds to an intracellular oxygen tension of about 0.2 mm (47).

Combined Fluorometer and Double‐Beam Spectrophotometer for Reflectance Measurements

An apparatus suitable for recordings of extracellular oxygen tension in terms of the degree of hemoglobin dissociation and intracellular oxidation‐reduction state in terms of the fluorescence level of mitochondrial and cytoplasmic reduced pyridine nucleotide has been constructed. A time sharing sequence permitting light and dark intervals for reflectance fluorometry and two flashes of different wavelengths for reflectance spectrophotometry operates at a repetition frequency of 60 cps. Fixed filters are used in fluorometry for excitation and emission and continuously variable interference filters are used in spectrophotometry.

Control of oxidation-reduction state of NADH in the liver of anesthetized rats

Abstract Simultaneous observations of fluorescence changes in the kidney and liver of the anesthetized rat are here interpreted in terms of oxidations and reductions of intracellular pyridine nucleotide. The pyridine nucleotide of the kidney only partially reduced in aerobiosis shows a large increase in reduction in anaerobiosis. In aerobiosis the level of reduction is sensitive to Amytal and it is concluded that the kidney is working under conditions of high energy drain in which phosphate and phosphate acceptor are in excess. The kidney shows a critical oxygen tension which corresponds to a small value of inspired air, probably less than 3 per cent. The oxidation-reduction status of NAD of the liver corresponds to that of a tissue in which the activity of oxidative phosphorylation sufficiently exceeds the energy demands upon the mitochondria so that the level of ADP and phosphate is sufficiently low to give a controlled state characterized by a high degree of reduction of NAD. The critical oxygen tension for the liver is high and corresponds roughly to about 5 per cent oxygen in the inspired air. In anoxia the kidney shows increased reduction of pyridine nucleotide while the liver shows a decreased reduction of pyridine nucleotide. Furthermore, in the oxygenation of the tissue following an interval of hypoxia, the liver mitochondria show increased reduction while those of the kidney show increased oxidation. A hypothesis which attempts to unify these apparently diverse conflicting observations is based upon the idea that the energy load in the kidney drops in hypoxia while that of the liver is constant.